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The vascular system of the femur and humerus was investigated in 17-24 wk human fetuses by scanning electron microscopy of corrosion casts. The number of nutrient foramina present in both bones ranged from 1 to 3 and the number of nutrient vessels associated with individual foramina also varied. The medullary arteries supplied both the bone cortex and marrow. There was no arterial supply to the shaft cortex from the periosteal side, where only capillaries were found to enter the bone. The metaphyses were supplied and drained by conspicuous vascular triads composed of an artery and 2 veins. In the marrow cavity, 2 morphologically different areas of the fine vascular network could be distinguished: diaphyseal sinusoids and metaphyseal capillaries forming a 'vascular besom' which was abruptly demarcated by the growth plate cartilage. The cortical microvascular bed was composed of capillaries and more numerous irregular sinusoids. The 2 main vascular systems, nutrient and periosteal, were interconnected not only via the network of cortical capillaries/sinusoids, but also by larger arteries and veins traversing the cortex. The features of the vascular system of human fetal long bones suggest its considerable functional flexibility and its capacity to modify blood flow patterns depending on circumstances.

In a computer simulation, growth of a capillary network is driven by a stochastic process on a planar hexagonal grid. Starting at a point source, the probabilities for the formation of new capillary elements depend on local biophysical knowledge. This knowledge is mainly derived from the flow theorem of Hagen-Poiseuille and the diameter exponent delta. The hexagonal grid is visualized as being supported by a cylinder or a sphere. An arterial tree results from the adaptive diameter augmentation, and is considered to have limited fractal properties. The dimension of its border, and the time course of growth and of blood pressure are compared with biological data from the chorioallantoic membrane (CAM) of incubated chicken eggs. The model is discussed in view of mechanosensitivity and cell-matrix interactions of endothelial cells, and CAM haemodynamics.

Purpose: The temporal and spatial sequence of development of laminar vascular plexuses was determined qualitatively and quantitatively in monkey and human retina., Methods: Histologic and cytochemical methods were used to study Macaca monkey eyes from fetal day 55 (F55d; birth = F168d) to 17 years, and human retina from fetal 21 weeks to adult., Results: In monkey retina, spindle-shaped, presumed vascular precursor, cells appear at F55d in the nerve fiber layer (NFL) adjacent to the optic nerve. The vascular plexuses in the NFL-ganglion cell layer appear first and form in the presence of spindle cells. Nerve fiber layer vessels extended radially to reach the temporal ora at F95d and nasal ora at F110d. The capillary plexus at the inner border of the inner nuclear layer (INL) appears at F120d near the optic disc, whereas the plexus at the outer INL border appears at F130d. Both reach their final position before birth. The INL plexuses form by endothelial budding from more vitread vessels in the absence of spindle cells. In the NFL, vessel growth to match retinal growth at the ora also involves endothelial budding. The growth rate of all plexuses was approximately 225 microns/day. The central fovea and the most peripheral retina adjacent to the ora serrata remained avascular throughout development. Differences between humans and monkeys include: Human vessels complete maturation after birth; human vessels reach the nasal ora earlier than the temporal ora; and spindle cells are more abundant and dispersed over a greater area within human NFL. Growth rates of human plexuses were comparable to those in monkeys., Conclusion: In both primates, deeper capillary plexuses form only by extension from existing vessels (angiogenesis). In the NFL, early vessel formation involves spindle precursor cells (vasculogenesis). The main difference between monkey and human in these processes is that the mature monkey vascular pattern is established well before birth.

Japanese quail embryos were used to examine paths of neural crest cell (NC) migration in relationship to the embryonic vasculature. Immunolabeling for NC, angioblasts and the extracellular matrix (ECM) glycoproteins fibronectin (FN), laminin (LN) and tenascin (TN) revealed several instances where spatiotemporal patterns of NC migration coincide with the embryonic vascular pattern and its associated ECM. An in vitro model for angiogenesis was modified to include NC, and associations with "capillary-like" endothelial cell structures were demonstrated. A working hypothesis is that the embryonic vasculature may, in specific instances, be used as a substratum for directed NC migration and that these interactions are mediated primarily through the adhesive interactions of FN. Some members of the TN family of glycoproteins, through their relatively non-adhesive properties, may act to help guide neural crest cells to the FN-rich blood vessel surface.

The neovascularization of the rabbit phallus at ages between prenatal days 15 and 21 was investigated by light- and electron microscopy, computer-aided light microscopic reconstruction, and immunocytochemistry. The phalli are embedded by an abundance of mesenchymal cells, which are in contact with the neighboring ones or with the endothelial lining of growing capillaries. They often form solid cell cords that eventually make contact with the growing capillaries. The computer-aided reconstruction of the serial light micrographs reveals that these cell cords are involved in connecting the adjacent capillaries. The incorporation of such mesenchymal cell projections into the endothelial lining, occasionally conjugated with simple attachment devices, is frequently observed by transmission and scanning electron microscopy. The contact areas between the mesenchymal and endothelial cells show immunoreactions of fibronectin. These results indicate the successive transformation of mesenchymal cells to endothelial cells of the growing capillaries. As endothelial cells of the growing capillaries show mitotic proliferation, such vasoformative mesenchymal cells seem to be involved in the acceleration of the neocapillarization of the rabbit phallus. Fibronectin actively produced in the mesenchymal cells may participate in their migration and the mechanical linkage with the endothelial cells.

The rate and pattern of growth as well as vessel ultrastructure of the area vasculosa were examined in the chick. The embryos were grown in shell-less culture after 3 d in ovo and staged according to Hamburger & Hamilton (1951) and the rate of increase in the diameter of the area vasculosa was measured. This revealed an increase in the area vasculosa diameter of 0.4 +/- 0.02 mm h-1 (n = 62) for embryos between stages 15 and 20. To determine the growth pattern of the sinus terminalis (the advancing edge of the area vasculosa), a marked length of the sinus was photographed at hourly intervals over a period of 9 h. It was found that this vessel grows by new vessels forming external to the sinus in the form of parallel plexuses, one of which then replaces the original sinus as the major route of bloodflow. Ultrastructurally the capillaries of the area vasculosa were simple tubes of endothelial cells, lacking a basement membrane. The endothelial cell cytoplasm contained only a few organelles, mainly mitochondria and rough endoplasmic reticulum. These findings indicate that the chick area vasculosa capillaries bear similar structural and growth characteristics to those associated with tumour angiogenesis and suggest that they may prove to be a useful model system for studying the factors involved in pathological angiogenesis.

The effect of vascular endothelial growth factor (VEGF165) on the chorioallantoic membrane (CAM) of 13-day-old chick embryos was studied. The factor was applied in doses of 0.5-4 micrograms for a period of up to 4 days. Macroscopical, histological and immunohistological studies were carried out. The localization of the factor was examined with an anti-VEGF antibody. The mitogenicity of VEGF165 and basic fibroblast growth factor (bFGF) were studied by means of the BrdU-anti-BrdU method. Furthermore, the effect of heparin alone and in combination with VEGF165 was investigated. VEGF165 specifically induces angiogenesis in doses of 0.5 microgram and more. A brush-like formation of blood vessels can be seen in the region of the precapillary vessels. Angiogenesis also takes place in the region of the capillaries and the venules. Histologically we found indications of sprouting as well as of intussusceptive capillary growth. The presence of the factor in the application area could be demonstrated with the anti-VEGF antibody for a period of 3 days. The factor is located in the chorionic epithelium and the intraepithelial capillaries. The BrdU-studies show that VEGF165 induces strong endothelial cell proliferation, whereas bFGF elicits fibrocyte proliferation and minor endothelial cell proliferation. Heparin induces squamous metaplasia of the chorionic and allantoic epithelium in combination with an aggregation of fibrocytes. We could not detect any enhancement of VEGF165 by heparin.

We investigated the development of human intrahepatic peribiliary capillary plexus by lectin-histochemistry and immunohistochemistry, using 40 fetal livers of various gestational ages, 52 postnatal livers of various ages, and 10 adult livers. In the earliest developmental stage of intrahepatic bile ducts ("the stage of ductal plate"), no vasculatures were found around the ductal plate, but progenitor vascular cells positive for von Willebrand factor, Ulex europaeus agglutinin I and succinylated wheat germ agglutinin appeared in the mesenchyme of the portal tracts. In "the stage of biliary cell migration into the mesenchyme," the progenitor vascular cells transformed into capillaries positive for von Willebrand factor, Ulex europaeus agglutinin I, and succinylated wheat germ agglutinin. In "the stage of bile duct formation," capillaries began to surround the bile ducts (immature peribiliary capillary plexus). The capillaries of the immature peribiliary capillary plexus proliferated and formed premature peribiliary capillary plexus around 40 wk of gestation in large bile ducts and around 6 wk after birth in small bile ducts. The former was composed of inner and outer layers, whereas the latter consisted of scattered capillaries without layer formation. The premature peribiliary capillary plexus continued to proliferate in the postnatal life and reached an adult and mature state around 15 yr. These data suggest that endothelial cells of capillaries of peribiliary capillary plexus derive from mesenchyme at the earliest stage of bile duct development and that the development and maturation of peribiliary capillary plexus progress parallel to those of the intrahepatic bile ducts.

This investigation was initiated with the intent of study capillary sprouting in the tadpole tail fin microcirculation of the African clawed frog, Xenopus laevis, using a combination of intravital video recordings and electron microscopy. The tadpoles were observed daily for periods up to one hour during one to two weeks. The capillary sprouts originated mostly from postcapillary venules, and within 24-48 h merged with other capillary sprouts, subsequently establishing a capillary loop with blood flow. As the circulatory patterns developed further, capillary regressions also occurred. As the electron microscope analyses of the capillary sprouts progressed, it became obvious that a thorough electron microscope study of the blood vessels of the tadpoles was required in order to explore structural characteristics of arterial and venous blood vessels. Thus, this article deals primarily with the general organization of the tadpole tail circulation, and the ultrastructural characteristics of the vascular walls. A subsequent article will deal with the role of endothelial cells, fibroblasts and pericytes in the process of capillary sprouting and regression, based on intravital recordings and electron microscope analyses.

The aim of our investigations was to test whether the chicken chorio-allantoic membrane (CAM) could be an adequate in vivo model for a new mode of capillary growth, originally described in the rat lung and termed intussusceptive microvascular growth. According to that concept the capillary system does not grow by sprouting of vessels, but expands by insertion of transcapillary tissue pillars or posts which form new intercapillary meshes. In the present study, we observed slender transcapillary tissue pillars with diameters around 1 microns in the CAM by in vivo microscopy, and analyzed their ultrastructure by transmission electron microscopic investigation of serial sections. The pillars corresponded in size to those previously described in rat lung microvasculature. On day 7, the pillar core contained endothelial-, endothelial-like cells and collagen fibers, and on day 12 additionally chorionic epithelial cells. As a hypothesis we propose that slender cytoplasmic extensions of endothelial cells, heavily interdigitated in the post area and often projecting into the vascular lumen, could initiate the first step of pillar formation, i.e., interconnect opposite capillary walls. During both stages of development endothelial-like cells were observed in close relationship with the pillars. These cells seem to be relevant for tissue post completion and growth, as they were found to invade the core of the pillars. From the localization of the interendothelial junctions in the post region, a certain similarity to the concept proposed for the lung can be found. The observations confirm that the CAM is a very suitable material for the in vivo investigation of intussusceptive capillary growth.

The expression of the cytoskeletal filaments vimentin and GFAP has been analyzed by immunocytochemical techniques in endothelial cells, pericytes, and astrocyte perivascular endfeet of microvessels of chicken optic tectum and cerebellum during embryonic development and in adulthood. Endothelial cells and pericytes were characterized by strong vimentin-immunoreactivity in both tectum and cerebellum only in early developmental stages (11-15 incubation days, i.d.). Astrocyte processes closely associated with the vessel wall were vimentin stained in the 11 i.d. cerebellum and vimentin-and GFAP-reactive in 15 i.d. tectum. These perivascular endfeet became GFAP-immuno-stained in the tectum and cerebellum by the 21st i.d. The results indicate that intermediate filament expression in the cells of the brain microvasculature is developmentally regulated, and suggest that the vimentin to GFAP transition in perivascular astrocytes parallels the vessel wall maturation.

Proliferative growth of the ventricular myocyte (cardiomyocyte) is primarily limited to fetal and early neonatal periods of development. In concert with the neonatal "transition" from proliferative to hypertrophic growth, ventricular remodeling of the nonmyocyte compartment is characterized by increased extracellular matrix synthesis/deposition and capillary angiogenesis. A role for locally generated and bioactive ventricular acidic fibroblast growth factor (aFGF) in these processes is proposed and substantiated by the following: 1) colocalization of aFGF peptide and fibroblast growth factor receptor (flg) transcripts to the developing fetal cardiomyocyte by immunohistochemistry, immunoelectron microscopy, and in situ hybridization, 2) continued localization of aFGF peptide and transcripts to the neonatal/mature cardiomyocyte, and 3) localization of flg immunoreactivity and transcripts to specific neonatal ventricular nonmuscle cell types. Specific ventricular cell types at distinct developmental stages appear to be responsive to ventricular myocyte-derived aFGF (myocytes in the fetal heart and nonmyocytes/endothelial cells in the neonatal heart). These data indicate that expression of aFGF and one of its receptors (flg) are most pronounced in the fetal to early neonatal ventricle, the presence of both suggesting an autocrine/paracrine growth regulatory function. As the animal matures, ventricular capillary angiogenesis may be facilitated by "release" of cardiomyocyte-derived fibroblast growth factors into the surrounding extracellular space/matrix functioning as a "paracrine" angiogenic stimuli. Therefore, the results of our study suggest that myocyte-derived aFGF may function to increase the fetal ventricular cardiomyocyte population in absolute number as well as to facilitate the subsequent increase in capillary angiogenesis that occurs during cardiomyocyte maturation and ventricular remodeling.

The occurrence of intraventricular hemorrhage (IVH) is a frequent problem in premature infants delivered between 24 (60%) and 32 weeks (80%) of gestation, a time during which the germinal matrix (GM) is prominent over the head of the caudate nucleus. Most IVH arises from the GM and it has been proposed that an important factor in the pathogenesis of IVH is a weakness of GM capillary walls due to deficient support by surrounding immature glial cells. The purpose of this study was to examine the glial-capillary interaction in the GM of fetal baboons sacrificed at 100 days (54%) gestation, a stage of GM development comparable to that during which human neonatal IVH occurs. Brains from a later gestational stage (162 days, 88%), after GM involution, were also examined. At 100 days of gestational age, the GM was prominent over the head of the caudate and contained vimentin positive, but not glial fibrillary acidic protein positive, radial glial cells which formed endfeet on capillaries in the region. Ultrastructurally, all the GM capillaries examined from this gestational time had complete, continuous endothelia marked by few pinocytotic vesicles and prominent tight junctions. The endothelial cells rested upon uninterrupted basement membranes which were contacted by clearly identifiable glial endfeet. These data show that GM capillaries have morphologies typical of CNS capillaries and suggest that the capillary immaturity within the GM is not a major contributing factor to IVH.

The spatial distribution of capillaries in the optic chiasma was studied quantitatively in 51 fetuses, 8 newborns and 4 babies to reveal that the capillary density was lower in the chiasmal median portion, which was therefore a weak point of blood supply. This finding may be a contributing factor for bitemporal hemianopsia due to neoplasms from the sella turcica.

Vascularization and expression of blood-brain barrier (bbb) associated morphological characteristics were studied by ultrastructural analysis during the embryonic development of the mouse CNS. At day 9 (E9) capillaries were only found in the perineural mesenchymal tissue exhibiting fenestrations and loosely attached pericytes. At E10 endothelial cells (EC) together with pericytes and other mesenchymal cells invaded the intraneural domain. The immigrating EC showed numerous protrusions and lost their fenestrations as soon as the new intraneural capillaries were formed. Intraneural EC showed prominent nuclei, many pinocytotic vesicles and junctional complexes which appeared tight in some places. These results indicate that from the first day of intraneural vascularization onwards, the morphological properties of the bbb are present in the early embryonic mouse cerebral cortex.

There is still some controversy on condylar cartilage growth capacity, whether it is similar or not to long bones growth cartilage. Recently it has been set that human tibial proximal growth decreases gradually throughout the second half of pregnancy period. The aim of this study is to determine whether changes are produced within mandibular condylar cartilage during fetal period are similar or not those that take place in growth platform. Lateral X-rays from the right half of the mandible of 19 human foetus between 18 and 41 weeks had been obtained. Total mandibular length, corpus length, ramus length and gonial angle were measured. A hystological condylar study was made, recording its thickness from sagittal sections of the central condylar area. Total mandibular length as well as corpus and ramus length progressively increased during studied period, existing high correlation between those parameters and gestational age and weight. Gonial angle decreases in studied period. Changes in condylar thickness were correlated neither with pregnancy age nor with fetal weight or X-rays mandibular parameters. These outcomes show that changes in mandibular condyle are not similar to those produced in tibial growth proximal platform and suggest condylar modifications could be more related to local factors than with general body growth mechanisms.

On cryostat sections of hearts of human embryos and fetuses aged 7 to 20 weeks, the length, density and internal surface (area) of capillaries were studied in surface unit of the ventricular myocardium using a semi-automatic apparatus MOP AM 03. The DPP IV and ALP-positive capillary segments were evaluated separately.

To investigate cardiovascular adaptation to chronic anemia we studied eight ovine fetuses made anemic by serial isovolemic hemorrhage and seven nonanemic controls. After 1 wk carotid arterial oxygen content was reduced to 1.6 +/- 0.2 ml/dl and hematocrit to 13.3 +/- 1.6% in anemic fetuses compared with 6.9 +/- 1.2 ml/dl and 32.4 +/- 3.9% in controls. Cardiac output was higher in the anemic group (753 +/- 102 vs. 490 +/- 66 ml.min-1.kg fetus-1) as stroke volume and heart rate both increased. Blood flow to the carcass, skin, kidneys, intestines, brain, and heart was increased. Vascular resistance fell in all tissues except the placenta. Central venous pressure, arterial pH, plasma total protein, and blood volume were not different although extravascular fluid accumulated in six of the anemic fetuses. The estimated capillary hydrostatic pressure was greater in anemic (7.6 +/- 1.8 mmHg) than control fetuses (5.0 +/- 1.5 mmHg) and the ratio of precapillary to postcapillary resistance was less. We conclude that reduction in the ratio of precapillary to postcapillary resistance in chronic fetal anemia increases blood flow, oxygen delivery, and capillary hydrostatic pressure.

Basement lamina and pericytes of growing blood microvessels were analyzed in the chick embryo optic tectum, from the 8th incubation day to hatching. Formation of the basement lamina and morphological changes of the pericytes take place in a short range of time, but late in the embryonic life, when also the blood brain barrier (bbb) devices are developing. The spatial and temporal coincidence between basement lamina formation, endothelium tight junction differentiation, and perivascular arrangement of the astrocytic glia, indicates that these events are correlated and corroborates the hypothesis that the glia needs an extracellular matrix to induce the junctional system maturation in the neural endothelia. Pericytes are irregular in shape during the early neural angiogenesis and smooth and flattened later, as the basement lamina synthesis is taking place; these cells represent a second line of barrier beyond the endothelium when the bbb is immature, owing to their phagocytic and digestive properties.

The microvascular endothelium of the chick wing bud at stages 22, 27, and 32 was evaluated by ultrastructural morphometry. The rationale for this study is based on the hypothesis that endothelial cells exhibit variation in structure and function during cytodifferentiation. The microvessels had a luminal diameter range such that they were classified as capillaries. The thin continuous endothelium was devoid of a basal lamina. The endothelium had a very small number of plasmalemmal vesicles; vacuoles were however present for all stages and in some cases were abundant. The temporal findings were that endothelial cell thickness increases, plasmalemmal vesicle densities decrease, and the densities of cytoplasmic vacuoles increase. The spatial results were that endothelial cells in proximal regions of the limb have a greater thickness, contain fewer vesicles and have more vacuoles than those in distal regions. In general, these results indicate that endothelial ultrastructural heterogeneity occurs within a 3 1/2 day time-span of wing bud development. The discussion considers the results with regard to recent reports on endothelial cell heterogeneity.

Morphometric changes in the rat lateral ventricle choroid plexus epithelium and endothelium and in the ventricular ependyma were studied between 16 days gestation and 30 days after birth, using stereological techniques. The epithelial apical surface density increased from 0.6 to 3.3 microns 2/microns 3 and the mitochondrial volume fraction from 3.2 to 7.6% during this period. The endothelial fenestrations increased from 0.05 to 0.39 micron-1. These changes may be related to postnatal increases in choroid plexus function. Morphological changes in basolateral surface density, cell height and nucleus and glycogen volume fraction have also been measured. The development of the lateral ventricle choroid plexus was qualitatively similar to the fourth ventricle plexus reported previously, but small quantitative differences occurred. The ventricular ependyma also showed a significant increase in mitochondrial volume fraction after birth, though to a lesser extent than the plexus epithelium. The total apical surface area of the choroid plexuses was estimated at 75 cm2 for 30-day-old rats. This figure, which takes into account the apical microvilli, is much greater than previous estimates and is similar to the surface area of the cerebral capillaries (155 cm2), and suggests that the choroid plexuses may play a more important role in the regulation of the brain microenvironment than previously thought.

Succession in formation of blood bed of the greater omentum has been studied in human fetuses (42), in premature children with the body mass of 900-2,500 g (108) and in mature children up to 3 years of age (48). Vasculogenesis in the organ of 12-week-old fetuses and in older ones is demonstrated as an ansiform growth of the vessels. In the vascular loop the afferent geniculum contains differential signs of the arterial, and the different one--the venous part of the blood bed. The top of the loop contains a growth bud and is a zone of growth; it ensures proliferation of the vascular loops and their spreading in the growing tissue mass in the organ. This principle of growth provides a ramified system of arteries and veins, reiterating the course and branching of each other. Against the background of the ansiform growth of the vessels capillary formation begins, at first connective, and then true capillaries are formed. Basing on the data obtained a conception of vasculogenesis in ontogenesis is formulated. Its main stages are--period of ansiform growth of the vessels and period of capillary formation. In the latter should be distinguished appearance of connective capillaries and formation of truely capillary network.

The development of the vasculature of the pars distalis of two strains of rat, Fischer 344 (F344) and Lewis (LEW), was followed in 16-day (16d) and 20-day (20d) fetuses, and in 1-day (1d), 5d, 20d, 50d, and 6-month-old females. No differences in the two strains were apparent in 16d fetuses; and the capillaries that were present were immature, i.e., tall, non-fenestrated endothelial cells, and were surrounded by poorly delineated pericapillary spaces. Immature capillaries also were predominant in 20d fetuses of both strains. Agranular folliculo-stellate cells were identifiable, projecting endfeet to the parenchymal basal lamina in 20d F344 fetuses, but not in LEW fetuses. Postnatally, the capillaries of LEW rats became progressively more thin-walled and fenestrated, and were surrounded by a pericapillary space that was well delimited by basal laminae at 20d. In 50d and 6-month LEW rats, capillaries were intact and surrounded by well-defined pericapillary spaces. By comparison in F344 rats, the capillaries remained more immature even in 50d rats and older. In addition, in F344 rats focal disruptions in endothelial cells and disruptions in parenchymal and capillary basal laminae were present in all postnatal stages, and a dramatic accumulation of plasma was evident within the pericapillary spaces at 20d. Endfeet processes of folliculo-stellate cells were abundant at the parenchymal basal lamina of 1d and 5d F344 neonates, but only rarely were identified in LEW neonates. Some activation of folliculo-stellate cells, i.e., increased numbers of lysosomes and dilated endoplasmic reticulum, was present in 50d F344 rats. Connective-tissue cells within the pericapillary space also were numerous and activated in F344 rats. Discrete gaps in the parenchymal basal lamina were evident subjacent to the folliculo-stellate cell endfeet in F344 rats but not in LEW rats. The vascular bed of F344 rats differs in its development from that of LEW rats. Characteristic of the F344 strain is a persistence of more immature capillaries, an inherent vascular fragility, and an activated state of folliculo-stellate cells.

The appearance of the basement membrane components laminin, heparan sulphate proteoglycan and Type IV collagen was investigated by indirect immunofluorescence during the onset of vascularisation of the neuroepithelium of mouse embryos (Days 10, 11 and 12 of development). All three basement membrane components could initially be seen in the embryonic CNS anlage along with the appearance of capillaries in the neuroepithelial cell layer. From this direct correlation between vascularisation and appearance of basement membrane components in the embryonic anlage one can assume that the basement membrane components laminin, heparan sulphate proteoglycan and collagen Type IV are involved in the initial vascularisation of the CNS anlage of 10 to 12 day old mouse embryos.

The migration of capillaries into mouse embryonic kidneys grafted on quail chorioallantoic membrane (CAM) was analyzed by two monoclonal antibodies against quail endothelial and haematopoietic cells. As shown by immunohistochemistry, the quail chorioallantoic vessels invaded the kidney explant. Initially, the capillaries were detected in the interstitial stroma and, soon thereafter, tightly adjacent to the branches of the ureteric bud. The induced mesenchymal cell condensates, the prospective nephric vesicles, were avascular, but when the early S-shaped body was formed, the capillaries invaded its lower crevice. Finally chimeric glomeruli consisting of mouse podocytes and quail endothelial cells, were formed and, contemporarily, the capillaries ceased to migrate. Within the endothelial-mesangial area of the chimeric glomeruli, all cells expressed the quail-type nuclear structure and were stained by the quail endothelial-specific antibodies. The pattern of migrating capillaries was compared to the distribution of the extracellular matrix (ECM) molecules by double staining with polyclonal antibodies against laminin or fibronectin, and monoclonal quail endothelial-specific antibodies. Initially, the capillaries migrated in a fibronectin-rich matrix, devoid of laminin, but when the epithelial kidney tubules formed, some capillaries attached to the newly formed epithelial basement membrane. At no stage were the capillaries seen to penetrate the epithelial basement membrane. The orderly branching of the ureteric bud, followed by the formation of nephrons and the shift in the ECM, might create pathways for an oriented capillary migration. The fibronectin-rich areas could be a scaffold for the capillary migration, and the attachment to the basement membranes a means for their cessation.

The development of the pituitary gland and its blood vessels is described in rat embryos (gestational day 12 through day 21) injected with India-ink via the umbilical vessels. The vascularization of all components of the pituitary gland develops from the surface network covering the prosencephalic vesicle. However, vascular connections exist between the prospective median eminence and the anterior pituitary gland in the earliest stages examined (day 12) but are not augmented by vessels from the stomatodeal roof until day 13. Primary portal veins appear initially on day 13, the vascularization of the pars distalis is visible on day 15. The "Mantelplexus" covering the floor of the diencephalon is discernible on day 16. Large-caliber portal veins appear immediately before birth, but otherwise there is no significant change in the vascular pattern during the last five days of gestation. The pars intermedia and the median eminence-pituitary stalk region remain avascular throughout embryonic life.

Twelve human fetuses with radial aplasia were dissected to evaluate the vasculature of the arm. Three types of vascular patterns were found. In type I there was a single midline superficial vessel with no radial or ulnar artery. This pattern occurred in four perfused twins (acardia) with twins reversed arterial perfusion sequence, one fetus with clinical trisomy 18, and one with an unknown disorder with multiple field defects. In type II there was absence of the radial artery with or without persistence of the embryonic median artery; the other vessels were normal. This pattern was seen in three fetuses with unknown disorders associated with multiple malformations. In type III the radial artery was present but had an abnormal course. This pattern was found in three fetuses with thrombocytopenia-absent radius syndrome. From these dissections and the results of animal experiments on the embryology of the vasculature, the following conclusions can be drawn. Vasculogenesis precedes differentiation of mesenchyme into muscle and bone. The embryonic capillary net determines the adult vascular anatomy of the limb. Failure to form, malformations, or disruption of the capillary net results in anomalies of the adult vasculature and may lead to musculoskeletal defects. Failure to chondrogenesis does not disturb the capillary net, and normal arteries will develop. Absence of the radial artery found in association with radial aplasia implies that there was abnormal vasculogenesis or disruption of developing vessels, or both. Either can lead to this congenital musculoskeletal defect.

Augmentation of cytoskeletons in fetal capillary endothelial cells is one of the well known pathologic changes in toxemic placentae. But we can find very few reports on these filaments in placentae, especially on their ontogeny. An ultrastructural study on human chorionic villi from 55 normal and 25 toxemic pregnancies, covering all periods of gestation, revealed that endothelial cells already had a small amount of intermediate filaments and microfilaments from 6 weeks' gestation and that they gradually increased as pregnancy progresses. In toxemic cases, intermediate filaments were markedly augmented, occasionally forming bundles, to occupy the cytoplasm of endothelial cells, so that there was often no space for cytoplasmic organelles. Intermediate filaments were predominant in thinner parts of endothelial cells; both intermediate filaments and microfilaments were easily recognized in thicker parts. These filaments seemed to determine the arrangement of organelles such as rough endoplasmic reticulum and mitochondria. Immunohistochemical and immunoelectron microscopic studies with monoclonal antibodies proved that endothelial cells contained vimentin and actin, not cytokeratin and desmin. In toxemic placentae, augmentation of intermediate filaments may disturb normal movements of endothelial cells, and may cause an increase in vascular resistance to placental microcirculation and intrauterine growth retardation in the fetus.

The developmental stage of the mouse embryo at which capillaries first occurred and their localisation in the neuroepithelium were investigated on serial sections of 9 and 10 days old embryos embedded in paraffin and Epon. In addition, areas of the neuroepithelium in which capillaries had been observed at the light microscopical level were investigated by electron microscopy carried out on embryos fixed with glutaraldehyde supplemented with tannic acid. In 5-7 microns serial sections of paraffin-embedded embryos, capillaries were initially seen in the CNS anlage at Theiler's Stage 14 (1972). At this stage, capillaries also occurred in the prosencephalon, the rhombencephalon and in the developing spinal cord. In 1 micron serial sections of resin-embedded embryos, capillaries could be identified in the neuroepithelium one stage earlier, i.e. at Stage 13. These very early capillaries were seen in the dorsal part of the lateral wall of the prosencephalon which later forms the diencephalon. At the ultrastructural level, those areas of the neuroepithelium in which leptomeningeal capillaries first started to spread into the neuroepithelium were characterised by the disintegration of the basement membrane of the neuroepithelium and the appearance of tannic acid-positive extracellular structures between the neuroepithelium and the adjacent leptomeningeal capillaries.

The ulstrastructural characteristics of the cerebellum and olfactory bulb capillaries were studied in 12 human specimen measuring 25-200 mm. Type I capillaries, whose lumina were wider than 8 mu, contained no pores of fenestrations. The basement membrane was sometimes discontinous. In that case, junctions without quintuple-layered zones could be observed. Such capillaries may correspond to primary vessels in which the sinusoid character has disappeared. Elsewhere, the basement membrane was continuous and the interendothelial junctions always contained quintuple-layered zones. Type II capillaries, whose lumina measured between 2 and 8 mu had basement membranes that were either discontinuous or continuous. They were identical to the immature capillaries described previously in numerous species including man. Type III caillaries, rarely observed, were characterized by the presence of several endothelial cells with abundant cytoplasm, which limited the mina from 0.5 --3 mu in diameters. They had continuous basement membranes. Type IV and V capillaries has small or non-patent lumina. The basement membrane was absent or rudimentary. These capillaries appeared to correspond to simple and complexendothelial sprouts. In Types II, III, V and most Type IV capillaries, the interendothelial junctions contained quintuple-layered zones. No mature capillaries were observed. The hypothetical pathways of development of cerebral capillaries and the degree of permeability of immature interendothelial junctions are discussed. The long duration of maturation in man probably accounts for the diversity of capillary formations observed as compared with animals. The co-existence over along period of time of several capillary types with probably varying permeability may play a significant role in the maturation of the blood-brain barrier.

The developing capillaries of the mouse neurohypophysis were studied in the electron microscope to elucidate the fine structural differentiation of the vascular component of the neuro-hemal contact zones in the external median eminence and the neural lobe. In the embryo the growth of the superficial net of the primary plexus of the hypophysial portal system is largely manifested by the presence of proliferation areas located within the capillary plexus covering the surface of the median eminence. Presumptive shallow capillary loops diverge from these ares in the first postnatal week. Differentiation of the capillary wall follows the pattern outlined for continuous capillaries. A few fenestrae appear in the endothelium of immature, superficial vessels at the 17th gestational day, increase in frequency during the following embryonic days, and occur regularly in the postnatal animal. In the neural lobe the internal capillaries proliferative by vascular sprouts emanating from the vessels on the surface of the gland. At the end of embryonic time an extensive net has developed, composed of capillaries with immature characteristics. Proliferation is largely finished by the end of the third postnatal week, when mature capillaries dominate the picture. Formation of attenuated, porous areas is a postnatal process, apart from single fenestrae appearing in the walls of a few immature capillaries in late fetal life. The structural possibilities for an onset of neurohypophysial function in the mouse is discussed.

It is not known why the coronary arteries almost always originate only from the right and left aortic sinuses of Valsalva, since the structure and conditions appear to be the same for all six sinuses of the embryonic great arteries. We sought a possible mechanical explanation for the phenomenon by studying the development of the coronary vasculature in 351 staged, serially sectioned human embryos of Carnegie stages 9 through 23 from the Carnegie Embryological Collection. A plexus of blind epicardial capillaries appears on the heart in Carnegie stage 14 or 15 and acquires a coronary sinus connection in stage 15, 16, or 17. The connection of the proximal coronary arteries to the aorta does not appear until stage 18. We found no histologic features of the cardiac nerves or any other component of the tissues to account for the consistent origin of coronary arteries from the right and left aortic sinuses of Valsalva. However, serial section reconstructions showed that the two sinuses where coronary arteries develop acquire a positive transverse curvature and a negative longitudinal curvature, i.e., a catenoidal or saddle-shaped configuration, before the appearance of the coronary arteries. The four noncoronary sinuses also have a positive transverse curvature, but longitudinally, in contrast, they have a positive curvature or are straight. The results suggest that the coronary arteries originate from those sinuses of Valsalva where wall tension is increased by a catenoidal configuration.

In the late prenatal stage, the rabbit corpora cavernosa penis (cavernous bodies) are occupied by a large number of mesenchymal cells actively proliferating by mitosis. Most profiles of the cavernous sinuses show growing capillaries consisting of immature endothelial cells and enclosing a narrow lumen. Such growing capillaries are often associated with the mesenchymal cells that contact with each other and eventually with the endothelial cells. Between 1 and 7 postnatal days, the capillary network develops extensively and the maturation of the capillaries results in a flattened endothelium with a continuous basal lamina. The mesenchymal cells are also associated with the helicine arterial sprouts at this time period. No endothelial mitotic figures were observed within the growing capillaries in pre- and postnatal specimens examined. Single pulse labeling with 3H-thymidine in the postnatal rabbits demonstrates that the labeled endothelial cells of the growing capillaries increase in number at 48 hours after injection compared with those at 24 hours, whereas the labeled mesenchymal cells inversely decrease in number. These findings indicate that the incorporation of the mesenchymal cells as vessel-forming cells into the growing capillaries accelerates the neovascularization in the rabbit cavernous bodies.

The formation of interrelations of the axons of neurosecretory cells and of ependyme cells with the capillaries of primary portal plexus in rats from the 14th day of embryogenesis till the 9th day of postnatal life was studied using the light and electron microscope methods. During the whole period under study, the basal processes of the ependyme cells reach the primary portal plexus of the capillaries. The terminals of the basal processes are usually separated from the endothelium of the capillaries by two basal membranes and enclosed pericapillary space. After the birth, some basal process penetrate in the pericapillary space and terminate on the endothelium. The surface of contact of the ependyme cell processes with the external basal membrane increases with the age, this being accompanied by the increase of pinocytotic activity. The neurosecretory axons are found in the median eminence already on the 14th day of embryogenesis, but by the 20th day only they reach the external basal membrane and penetrate sometimes in the pericapillary space. After the birth, the number of axons reaching the external basal membrane and the surface of contact between them increase gradually with, apparently, a concomitant intensification of the transport of neurohormones in the portal circulatory system of the hypophysial-hypothalamic complex.

The data on formation and construction principles of the blood supply system in the terminal sensitive apparatuses of the human forearm muscles during embryogenesis and early postnatal period are presented. During the intrauterine development, the formation of the vascular-receptor relations takes place in the sequence corresponding to the anlage of the receptors. The first blood vessels appear in the area of the nonencapsulated sensitive terminals and only after in the encapsulated receptors. The latter have their own blood supply system which is formed by the vessels of the intramuscular neural fasciculi and the surrounding muscular tissue. Around the sensitive terminals thick microvascular networks are formed, they often tightly adhere the external receptor capsule. The intrareceptor capillaries are situated in the mass of the neural terminals and sometimes form capillary loops in them.

The development of the intracerebral capillaries of the telencephalon was studied by the combined techniques of microangiography and histology in embryos and fetuses of cattle and pigs. In the telencephalic colliculi ganglionares, vascularization starts during the 4th week of gestation and leads to the formation of a peripheral and a central arterial system separated by the internal capsule. Capillary development within the pallium occurs 3 weeks later and consists in the formation of longitudinal arteries, concentrically arranged rings of deviations and bifurcations, and a periventricular capillary network.

The elaboration of the basal lamina of telencephalic capillaries has been studied by autoradiography in chick embryos from day 16 to 19 of incubation. The cerebral vascular basal lamina is actively synthesized during the period of these stages of development. The animals were sacrificed 2, 15, 40 and 180 min after intravenous injection of 3H-proline. The hydroxylation of proline to hydroxyproline in the "secretory" cells permits their identification. Our observations suggest that endothelial cells, pericytes and perhaps glial cells participate in this elaboration. The various stages of the synthesis are performed sequentially in the ergastoplasm and then in the Golgi apparatus before the secretory material is released by exocytosis.

The characteristics of the germinal matrix vasculature were studied in the developing fetal brain using immunocytochemical methods. A preliminary comparative immunocytochemical study was made on six fetal brains to compare endothelial staining by Ulex europaeus I lectin with that of antibody to Factor VIII related antigen. Ulex was found to stain germinal layer vessels better than Factor VIII related antigen. Subsequently, the germinal layers of a further 15 fetal and preterm infant brains ranging from 13 to 35 weeks' gestation were stained with Ulex europaeus I to demonstrate the vasculature. With increasing gestation, there was a gradual increase in vessel density, particularly of capillaries. This was not a uniform process. A plexus of capillaries was prominent immediately beneath the ependyma while the more central parts of the germinal matrix contained fewer, but often larger diameter, vessels. The variation in vessel density which was a feature of the later gestation brains may have implications for local blood flow and may be a factor in haemorrhage at this site.

The prenatal development of the intracerebral capillaries within the brain stem was systematically examined by a special microangiographic technique in 69 embryos and fetuses of cattle and pigs between 4 and 17 weeks of gestation. The microradiograms were compared with the corresponding histological slices. The different developmental stages and morphological patterns of vascularization are related to the local stage of maturation, histological structure and metabolic activity of the nervous tissue.

The electronmicroscopic control of the capillaries of the brain vesicles in 5--6-day-old chick embryos revealed an intimate contact between capillary walls and neuroblasts, and on the other hand the presence of a functional endoplasmic reticulum in the endothelial cells. These are only very few vacuoles for protein transport in the cytoplasm of endothelial cells.